The present invention broadly is in the field of medical applications, and, more specifically, relates to particle analysis with electronic, optical or mechanical means and, in particular, is concerned with the field of analysis of blood particles.
It is extremely problematic to analyse a mixture of particles of different size depending upon the distribution of the prevailing classes. What is meant herein by the expression distribution as used in this disclosure, is a discrete distribution of body volume and, in the narrower sense of a histogram, the frequency of the body volumes. In the statistical sense the histogram or the distribution is the probability density that a feature or characteristic, as to its value, will lie within a predetermined interval. The characteristics or features can be of different nature, for instance physical, chemical, morphological and others. If interest is only expressed in the particle count of a certain class, which ideally is present isolated from the distribution of other classes, then the analysis is unambiguous. Prevailing errors usually are attributable to the employed equipment and the accuracy is only limited by the signal-to-noise ratio inherent for the employed system. However, as soon as there are present mixed distributions, wherein, for instance, the same size particles of different particle species or classes belong to a respective inherent distribution density, then there are required criteria for discriminating between the overlapping distributions.
If for such a mixed distribution it is known that all classes are normally distributed, then, for instance, the finite number of classes can be determined according to a method disclosed by G. Doetsch. Yet, practice of this method is extremely cumbersome and presupposes that the mixed distribution is already known through knowledge of the mean value and the variance. However, with the here interesting fields of application there is usually only known an empirical distribution. In this case, for instance, the probability paper furnishes a raw method or technique in order to determine the one mixed distribution of exactly two classes by appropriately trying the possibly basically prevailing normal distribution.
With the heretofore known particle analysis equipment there are employed the human efforts of the operator in order to separate certain ranges of a mixed distribution of the body volume. The operator evaluates the distribution spectrum or a function derived therefrom, which spectrum has been obtained in a not here further described manner, and based upon criteria which the operator selects determines a "separation threshold", below which, for instance, the particles are allocated in accordance with their size to one class and above which the particles are then allocated to the other class. The once selected threshold is impressed upon the system, the particle analyser then only detects, for instance, the signals related to the particles which are below or above this threshold. Assuming the signals below the threshold are predicated upon spurious particles, the signals above the threshold upon particles which must be analysed, then the set threshold constitutes a discriminator for spurious and useful signals. If there are measured distributions of a number of size classes, then there must be selected a correspondingly greater number of separation thresholds which must be impressed upon the measuring system, provided that the individual classes are satisfactorily separated in order to be even able to detect a mixed distribution. This also is true for a bimodal mixed distribution.
Now if a particle analyser is employed for a special purpose, in other words for a limited field of application, say, for blood particle analysis, then in a great many types of equipment the thresholds are fixed in order to separate the particles which should be counted or measured from the particles which should not be incorporated into the measurement.
The setting of separation thresholds in a multimodal distribution curve, in the first instance leads to truncated distributions. The degree of truncation has a direct influence upon the integral over the distribution density, for instance upon the result of a count, and determines, usually dominantly, its accuracy. This is only valid if one stays with the truncated distribution without adequately correcting the same. If the distribution curve changes at the region of a fixed threshold, then by virture of the increasing or decreasing truncation of the distribution to be analysed there is also altered the result of the analysis. While with prior faulty placement of the separation threshold it is possible for the result to become more accurate, normally however the opposite is true; the obtained result becomes poorer because the threshold previously usually was optimumly set. If, for instance, a particle analyser is designed for volume distribution analysis and for counting erythrocytes in human blood, that is to say, all of the sampled signals emanating from particles of a predetermined particle size interval should contribute to the measurement and a separation threshold should eliminate from the measurement those signals predicated upon artifacts, in other words, particles which are not erythrocytes, then this analyser, apart from possible exceptions, cannot be used, without correction of the threshold, for the counting of erythrocytes in animal blood. If the signal-to-noise ratio of the particle analyser is insufficient, then the physiologically possible variation range of the cell sizes in human blood already requires an individual accommodation of the threshold to each individual blood sample. Such analyser cannot be used at all for the analysis of just any random particles.
There will be clearly recognized from the foregoing the extremely narrow range of application with respect to a distribution function and, additionally, with insufficient signal-to-noise ratio the critical behaviour of a particle analyser with fixedly set separation thresholds.
System designs have been proposed wherein particle analysers are structured in such a manner that the operator, as required, can set the separation threshold or thresholds with the aid of a device mounted externally of the equipment. What previously was the task of an operator who was specially trained, now must be accomplished in equally exact and good quality by the particular random user of the equipment. The so-called setting or adjustment instructions should enable positive "setting" of a desired separation threshold by the user, without such manipulations falsifying the analysis results. Such setting instructions frequently are very simple, but, on the other hand, performance thereof is difficult and unreliable.
Thus, the threshold positioning or setting with the aid of an oscilloscope, where there are visible the particle signals and can be separated by varying a discriminator or threshold, delivers poorly reproducible values. Another recommended procedure requires the determination of a summation distribution curve. This is obtained by plotting counting results as a function of the threshold position. In the ideal case there is formed a horizontal segment, the so-called plateau, on the basis of which there can be set the threshold. The less the segment or plateau deviates from the horizontal and the greater its range, that much greater is the signal-to-noise ratio of the analyser. In the practical field of blood particle analysis the plateau, however, does not have any horizontal section and is also narrowly limited in range.
Positioning of the threshold on the basis of the determined curve is unreliable. In addition thereto, there must be considered the quite appreciable expenditure in time for the determination of the summation distribution curve, considering the fact that it must be periodically plotted and separately for erythrocytes and leukocytes. Additionally, the cell suspensions which are to be analysed are frequently unstable, something not known to many users. Consequently, the summation distribution curve is falsified and the threshold positioning based thereon is questionable. A possible solution from this dilemma is to improve the signal-to-noise ratio of the analysis system; through the use of sensors and a greater amount of electronic hardware it is possible to obtain a sub-critical threshold positioning. A further possibility is an adaptive threshold accommodation or setting.
Such threshold accommodation or setting is known from French Pat. No. 2,097,763. This patent discloses how to determine the transition between two overlapping distributions of signals. There are described means, with the aid of which there can be determined whether and in which channel of a histogram the stored frequency is smaller than the frequency stored in both directly neighboring channels. This known system serves for the determination of a local brightness extreme in a brightness field by finding and recognizing a characteristic object in a viewing field, wherein it is mentioned that the object can be a blood particle. The procedure which is followed is such that, after storing a complete histogram the counters of the individual channels are synchronously indexed or incremented until they are filled and in this state remain blocked. There is then determined which counter was the last to be filled. That is the counter of the channel where there is located the sought-for extreme. As will be apparent, with this procedure the histogram is extinguished, and therefore there is no longer possible an analysis of the signal distribution. In fact with this known teaching the procedures are carried out in two steps: in a first step there is determined, during the recording of a first histogram, the extreme and a threshold is set, then during a second step the histogram is again recorded and with the aid of the now set threshold evaluated. This procedure, where the first predetermined histogram is extinguished, is only capable of being used with those types of measurements where there is sampled an image which does not appreciably change as a function of time, whether such be the brightness image of a target position finder or homing device for navigation purposes, whether such be the image of a blood particle in the field of a microscope. The described procedure is not usable if there is not available an image which does not change as a function of time, for instance, if the signals are not brightness signals, rather designate the size of a particle in a particle suspension. In a particle analyser for use in a medical laboratory the histogram is formed rather slowly. With series tests it is important to maintain as short as possible the duration of each individual analysis, and therefore it is undersirable to perform each analysis twice (once in order to set a threshold, the second time in order to evaluate the histogram). Additionally, when proceeding according to the teachings of the aforementioned French Pat. No. 2,097,763 there is required twice the quantity of the particle suspension which is to be analysed than with a conventional particle analysis. This is prone to incur criticism by the potential customer of the analysis system.